GSM6932045: CD45-Ter119- cells from the bone marrow of PyMT-N tumor-resected mice, 28 days after resection; Mus musculus; RNA-Seq
To isolate the BM niche and HSPC cells, the muscle tissue on the tibia and femur were completely removed, then flushed out the BM cells. The remaining bone were cut and crashed into fragments and digest in DMEM containing 1 mg/mL Collagenase 1, 1 mg/mL Collagenase 2 , 4 mg/mL Dispase 2 , 0.1 mg/mL DNase 1 , 1 mg/mL BSA, 1% HEPES and antibiotics for 45 min at 37°C. Flushed BM cells also incubate in the same digest buffer for 15 min at 37°C. After digestion, bone cells and BM cells are filtered through a 70 µm strainer and resuspended in RBC lysis buffer, then combined into one sample (per mouse) for subsequent step analysis. Lin-cKit+ or CD45-Ter119- from the same batch of experiment were sorted into single-cell suspension and immediately submitted to the Single Cell Genomics Core at Baylor College of Medicine. The single-cell gene expression library was prepared according to the Chromium Single Cell Gene Expression 3v3.1 kit (10x Genomics).